影响PVDF膜对蛋白吸附效率的主要因素

发布时间：2018-03-19 06:52

本文选题：抗体纯化　切入点：PVDF膜　出处：《氨基酸和生物资源》2015年03期 　论文类型：期刊论文

【摘要】：通过结合有抗原的PVDF膜进行亲和纯化是小规模快速获取特异性抗体的一种重要手段。该方法的关键步骤之一是将抗原蛋白有效地吸附到PVDF膜上。尽管人们广泛使用PVDF膜亲和纯化抗体,但对影响PVDF膜吸附抗原蛋白的因素至今仍缺乏系统研究。以BSA为模型蛋白,检测在不同蛋白浓度、不同孵育时间、不同溶质溶液和不同pH值的情况下,PVDF膜吸附蛋白的效率。当蛋白溶液为PBS时,1cm2 PVDF膜与2mg·mL-1 BSA溶液孵育1h可达到其最大吸附率。常见的蛋白质缓冲液组分对PVDF膜的蛋白吸附率影响不一。其中,咪唑、Tris-HCl、NaCl、KCl在广泛的浓度范围内对PVDF膜吸附蛋白没有明显影响,而较高浓度的SDS、Triton-X 100、Tween-20、乙二醇、甘油、盐酸胍、尿素等可显著降低PVDF膜对蛋白的吸附效率,在pH为6.5~9.5的Tris-HCl中,随着pH的增高而吸附效率降低。
[Abstract]:Affinity purification with antigen-bound PVDF membrane is an important means to obtain specific antibody on a small scale. One of the key steps of this method is to effectively adsorb antigen protein onto PVDF membrane. The antibody was purified by PVDF membrane affinity. However, the factors affecting the adsorption of antigenic proteins on PVDF membranes have not been systematically studied. BSA was used as a model protein to detect different protein concentrations and incubation time. Under different solute solution and different pH value, the adsorption efficiency of membrane protein was obtained. When the protein solution was PBS, the maximum adsorption rate could be achieved when the membrane was incubated with 2 mg 路mL-1 BSA solution for 1 h. The common protein buffer components could reach the maximum adsorption rate on the eggs of PVDF membrane. The effect of white adsorption rate is different. Imidazolium Tris-HCln NaCl-KCl had no significant effect on PVDF membrane adsorption protein in a wide range of concentrations, while higher concentrations of SDSN Triton-X 100 Tween-20, ethylene glycol, glycerol, guanidine hydrochloride, urea and so on could significantly reduce the adsorption efficiency of PVDF membrane on the protein, and the adsorption efficiency of the membrane protein was significantly decreased when the pH was 6.59.5 Tris-HCl. The adsorption efficiency decreased with the increase of pH.
【作者单位】：华中农业大学农业微生物学国家重点实验室;
【基金】：中央高校基本科研业务费专项资金(2014PY003)
【分类号】：R392

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